Tetrahydrocannabinol (Delta-9-THC) is the major psychologically active component in the Cannabis plant (marijuana). Administration or use of marijuana or other products of the Cannabis plant, such as hashish and hash oil, can be detected through the analysis of biological fluids, such as blood or urine, for the parent compound delta-9-THC or its major urinary metabolite, namely, 11-nor-delta-9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH). Many different methods have been developed for the analysis of these compounds. Immunoassays are the most widely used today as discussed in the following publications:
Cook, C. E.; Radioimmunoassay of cannabinoids, IN: Cannabinoid Analysis in Physiological Fluids, ACS Symposium Series 98, Vinson, J. A., Ed., American Chemical Society, Washington, D.C., 137-154, 1979.
Teale, J. D.; Forman, E. J.; King, L. J.; and Marks, V.; Radioimmunoassay of cannabinoids in blood and urine, Lancet, 2:553-555, 1974.
Chase, A. R., Kelley, P. R., Taunton Rigby, A.; Jones, R. T.; and Harwood, T.; Quantitation of cannabinoids in biological fluids by radioimmunoassay, IN: Cannabinoid Assays in Humans, NIDA Research Monograph, No. 7, Willette, R. E., Ed., U.S. Department of Health, Education, and Welfare, Washington, D.C., 1-9, 1976.
Jones, A. B.; ElSohly, H. N.; and ElSohly, M. A.; Analysis of the major metabolites of delta-9-tetrahydrocannabinol in urine, V. Cross reactivity of selected compounds in a radioimmunoassay, J. Anal. Toxicol., 8:252-254, 1984.
Rowley, G. L.; Armstrong, T. A.; Crowl, C. P.; Eimstad, W. M.; Hu, W. M.; Kam, J. K.; Rodgers, R.; Ronald, R. C.; Rubenstein, K. E.; Sheldon, B. G.; and Ullman, E. F.; Determination of THC and its metabolites by EMIT homogenous enzyme immunoassay: A summary report, IN: Cannabinoid Assays in Humans, NIDA Research Monograph, No. 7, Willette, R. E., Ed., U.S. Department of Health, Education, and Welfare, Washington, D.C., 28-32, 1976.
Rodgers, R.; Crowl, C. P.; Eimstad, W. M.; Hu, M. W.; Kam, J. K.; Ronald, R. C.; Rowley, G. L.; and Ullman, E. F.; Homogenous enzyme immunoassay for cannabinoids in urine, Clin. Chem., 24:95-100, 1978.
However, in the performance of the above noted methods, especially in the forensic setting, it is recommended that they be conducted in conjunction with a confirmatory test based upon a different principle. The confirmatory test found most acceptable in the scientific community is gas chromatography/mass spectrometry (GC/MS). GC/MS is used not only to confirm the presence of the drug in the biological specimen, but also to quantitate the amount of the drug, as, for example, set forth in the following publications:
Nordqvist, M.; Lindgren, J. E.; and Augurell, S.; A method for the identification of acid metabolites of tetrahydrocannabinol (THC) by massfragmentography, IN: Cannabinoid Assays in Humans, NIDA Research Monograph, No. 7, Willette, R. E., Ed., U.S. Department of Health, Education, and Welfare, Washington, D.C., 64-69, 1976.
Karlsson, L.; Jonsson, J.; Aberg, K.; and Roos, C.; Determination of delta-9-tetrahydrocannabinol-11-oic acid in urine as its pentafluoropropyl-, pentafluoropropionyl derivative by GC/MS utilizing negative ion chemical ionization, J. Anal. Toxicol., 7:198-202, 1983.
Foltz, R. L.; McGinnis, K. M.; and Chinn, D. M.; Quantitative measurement of delta-9-tetrahydrocannabinol and two major metabolites in physiological specimens using capillary column gas chromatography negative ion chemical ionization mass spectrometry, Biomed. Mass Spectrom., 10:316-323, 1983.
ElSohly, M. A.; ElSohly, H. N.; Stanford, D. F.; Evans, M. G.; and Jones, A. B.; Analysis of human urine for 11-nor-delta-9-tetrahydrocannabinol-9-carboxylic acid: A comparison between HPLC, GC/ECD, GC/FID, and GC/MS methods, IN: Marijuana '84 Proceedings of the Oxford Symposium on Cannabis, Harvey, D. J., Ed., IRL Press, Oxtord, England, 137-146, 1985.
Internal standards most suitable for the GC/MS analysis are the deuterated derivatives of the drugs to be analyzed. The deuterated compound currently available for the analysis of THC-COOH is the trideuterated derivative, d.sub.3 -delta-9-THC-COOH. However, the employment of the trideuterated delta-9-THC-COOH presents several problems. First, this material is rather expensive and is not readily available. Second, the base peak in the mass spectrum of the deuterated standard, most commonly used as the methyl derivative, is at m/z 316 as compared to 313 for the natural metabolite. There is, however, a peak in the mass spectrum of the natural metabolite at m/z 316 as a result of a different fragmentation mechanism (through loss of --(CH.sub.2)4 from the side chain) which represents 4-5% of the base peak. Because the natural and deuterated acids are not chromatographically separable, this contribution to the 316 ion of the internal standard (I.S.) from the natural metabolite disturbs the ion ratios for the I.S. and adversely affects the quantitation. This is particularly true at higher concentrations of the natural metabolite.
The subject of this invention is the development of new internal standards which have essentially the same partitioning, chromatographic, and derivatizing behavior as do the compounds to be analyzed. Examples of these compounds include (a) hexadeutero-11-nor-delta-8-tetra-hydrocannabinol-9-carboxylic acid (d.sub.6 -delta-8-THC-COOH) for use as a internal standard for the analysis of THC-COOH, (b) nonadeutero-delta-9-tetrahydrocannabinol (d.sub.9 -delta-9-THC), (c) nonadeutero-delta-8-tetrahydrocannabinol (d.sub.9 -delta-8-THC), and (d) hexadeutero-delta-8-tetrahydrocannabinol (d.sub.6 -delta-8-THC). The latter three compounds have been developed for use as internal standards for the analysis of delta-9-THC in biological fluids.